Draft genome of neotropical Bacillus thuringiensis UFT038 and its potential against lepidopteran soybean pests.

Bacillus thuringiensis (Bt) is known for its Cry and Vip3A pesticidal proteins with high selectivity to target pests. Here, we assessed the potential of a novel neotropical Bt strain (UFT038) against six lepidopteran pests, including two Cry-resistant populations of fall armyworm, Spodoptera frugiperda. We also sequenced and analyzed the genome of Bt UFT038 to identify genes involved in insecticidal activities or encoding other virulence factors. In toxicological bioassays, Bt UFT038 killed and inhibited the neonate growth in a concentration-dependent manner. Bt UFT038 and HD-1 were equally toxic against S. cosmioides, S. frugiperda (S_Bt and R_Cry1 + 2Ab populations), Helicoverpa zea, and H. armigera. However, larval growth inhibition results indicated that Bt UFT038 was more toxic than HD-1 to S. cosmioides, while HD-1 was more active against Chrysodeixis includens. The draft genome of Bt UFT038 showed the cry1Aa8, cry1Ac11, cry1Ia44, cry2Aa9, cry2Ab35, and vip3Af5 genes. Besides this, genes encoding the virulence factors (inhA, plcA, piplC, sph, and chi1-2) and toxins (alo, cytK, hlyIII, hblA-D, and nheA-C) were also identified. Collectively, our findings reveal the potential of the Bt UFT038 strain as a source of insecticidal genes against lepidopteran pests, including S. cosmioides and S. frugiperda.

Novel Lactone-Based Insecticides and Drosophila suzukii Management: Synthesis, Potential Action Mechanisms and Selectivity for Non-Target Parasitoids.

Drosophila suzukii, an invasive insect pest, poses a significant threat to various fruit crops. The use of broad-spectrum insecticides to control this pest can reduce the effectiveness of biological control agents, such as the parasitoid Trichopria anastrephae. Here, we evaluated the toxicity of newly synthesized lactone derivatives on D. suzukii and their selectivity towards T. anastrephae. We used in silico approaches to identify potential targets from the most promising molecules in the D. suzukii nervous system and to understand potential differences in susceptibilities between D. suzukii and its parasitoid. Of the nine molecules tested, (rac)-8 and compound 4 demonstrated efficacy against the fly. Exposure to the estimated LC90 of (rac)-8 and compound 4 resulted in a mortality rate of less than 20% for T. anastrephae without impairing the parasitoid's functional parasitism. The in silico predictions suggest that (rac)-8 and compound 4 target gamma amino butyric acid (GABA) receptors and transient receptor potential (TRP) channels of D. suzukii. However, only the reduced interaction with TRP channels in T. anastrephae demonstrated a potential reason for the selectivity of these compounds on the parasitoid. Our findings suggest the potential for integrating (rac)-8 and compound 4 into D. suzukii management practices.

Methanolic Extracts of Chiococca alba in Aedes aegypti Biorational Management: Larvicidal and Repellent Potential, and Selectivity against Non-Target Organisms.

The use of formulations containing botanical products for controlling insects that vector human and animal diseases has increased in recent years. Plant extracts seem to offer fewer risks to the environment and to human health without reducing the application strategy's efficacy when compared to synthetic and conventional insecticides and repellents. Here, we evaluated the potential of extracts obtained from caninana, Chiococca alba (L.) Hitchc. (Rubiaceae), plants as a tool to be integrated into the management of Aedes aegypti, one of the principal vectors for the transmission of arborviruses in humans. We assessed the larvicidal and repellence performance against adult mosquitoes and evaluated the potential undesired effects of the extracts on non-target organisms. We assessed the susceptibility and predatory abilities of the nymphs of Belostoma anurum, a naturally occurring mosquito larva predator, and evaluated the C. alba extract's cytotoxic effects in mammalian cell lines. Our chromatographic analysis revealed 18 compounds, including rutin, naringin, myricetin, morin, and quercetin. The methanolic extracts of C. alba showed larvicidal (LC50 = 82 (72-94) mg/mL) activity without killing or affecting the abilities of B. anurum to prey upon mosquito larvae. Our in silico predictions revealed the molecular interactions between rutin and the AeagOBP1 receptor to be one possible mechanism for the repellent potential recorded for formulations containing C. alba extracts. Low cytotoxicity against mammalian cell lines reinforces the selectivity of C. alba extracts. Collectively, our findings highlight the potential of C. alba and one of its constituents (rutin) as alternative tools to be integrated into the management of A. aegypti mosquitoes.

Identification of potential new mosquito-associated viruses of adult Aedes aegypti mosquitoes from Tocantins state, Brazil.

Medically important arboviruses such as dengue virus (DENV), Zika virus (ZIKV), and chikungunya virus (CHIKV) are primarily transmitted by the globally distributed mosquito Aedes aegypti. Increasing evidence suggests that the transmission of some viruses can be influenced by mosquito-specific and mosquito-borne viruses. Advancements in high-throughput sequencing (HTS) and bioinformatics have expanded our knowledge on the richness of viruses harbored by mosquitoes. HTS was used to characterize the presence of virus sequences in wild-caught adult Ae. aegypti from Tocantins (TO) state, Brazil. Samples of mosquitoes were collected in four cities of Tocantins state and submitted to RNA isolation, followed by sequencing at an Illumina HiSeq platform. Our results showed initially by Krona the presence of 3% of the sequenced reads belonging to the viral database. After further analysis, the virus sequences were found to have homology to two viral families found in insects Phenuiviridae and Metaviridae. Three possible viral strains including putative new viruses were detected and named Phasi Charoen-like phasivirus isolate To-1 (PCLV To-1), Aedes aegypti To virus 1 (AAToV1), and Aedes aegypti To virus 2 (AAToV2). The results presented in this work contribute to the growing knowledge about the diversity of viruses in mosquitoes and might be useful for future studies on the interaction between insect-specific viruses and arboviruses.

Disentangling the ecotoxicological selectivity of clove essential oil against aphids and non-target ladybeetles.

The plant-based biopesticides have been proposed as insect pest control tools that seem to be safer for the environment and human health when compared to synthetic conventional molecules. However, such assumptions are generally made without considering the absence of detrimental effects on sublethally-exposed non-target organisms or showing the physiological basis of the selective action of such botanical products. Thus, by using in silico-based and in vivo toxicological approaches, the present investigation aimed to disentangle the ecotoxicological selectivity of clove, Syzygium aromaticum, essential oil against the aphid Rhopalosiphum maidis and the non-target ladybeetle, Coleomegilla maculata. We also investigated whether the sublethal exposure to clove essential oil would affect the locomotory and predatory abilities of C. maculata. We found that the clove essential oil concentration estimated to kill 95% (LC95: 0.17 µL/cm2) of the aphids was lethal to <18% of C. maculata. Indeed, our in silico results reinforced such differential susceptibility, as it predicted that eugenol and ß-caryophyllene (i.e., the clove essential oil major components) bound to three potential molecular targets (i.e., transient receptor potential (TRP) channels, octopamine, and gamma-aminobutyric acid (GABA) receptors) of the aphids but only to the octopamine receptors of the ladybeetles. Additionally, the ladybeetles that were exposure to the clove essential oil exhibited unaffected abilities to locomote and to prey upon R. maidis aphids when compared to unexposed ladybeetles. Thus, by displaying lower toxicity against the ladybeetles, the clove essential oil represents a safer alternative tool to be integrated into programs aiming to manage aphid infestations.

Comparative genomic analysis and mosquito larvicidal activity of four Bacillus thuringiensis serovar israelensis strains.

Bacillus thuringiensis serovar israelensis (Bti) is used to control insect vectors of human and animal diseases. In the present study, the toxicity of four strains of Bti, named T0124, T0131, T0137, and T0139, toward Aedes aegypti and Culex quinquefasciatus larvae was analyzed. The T0131 strain showed the highest larvicidal activity against A. aegypti (LC50 = 0.015 µg/ml) and C. quinquefasciatus larvae (LC50 = 0.035 µg/ml) when compared to the other strains. Furthermore, the genomic sequences of the four strains were obtained and compared. These Bti strains had chromosomes sizes of approximately 5.4 Mb with GC contents of ~35% and 5472-5477 putative coding regions. Three small plasmids (5.4, 6.8, and 7.6 kb) and three large plasmids (127, 235, and 359 kb) were found in the extrachromosomal content of all four strains. The SNP-based phylogeny revealed close relationship among isolates from this study and other Bti isolates, and SNPs analysis of the plasmids 127 kb did not reveal any mutations in δ-endotoxins genes. This newly acquired sequence data for these Bti strains may be useful in the search for novel insecticidal toxins to improve existing ones or develop new strategies for the biological control of important insect vectors of human and animal diseases.

Essential oil from Negramina (Siparuna guianensis) plants controls aphids without impairing survival and predatory abilities of non-target ladybeetles.

Plant essential oils are regarded as interesting alternative tools to be integrated into the management of pest insects. However, as they generally consist of mixtures of numerous molecules, the physiological basis for their action is unresolved. Here, we evaluated the effects of essential oil of the Neotropical plant Siparuna guianensis Aubl., commonly known as Negramina, against an important pest insect: the green peach aphid Myzus persicae (Sulzer), and also in two non-target natural enemies: the ladybeetle predators Coleomegilla maculata (DeGeer) and Eriopis connexa (Germar). In addition, we conducted a computational docking analysis for predicting the physical interactions between the two Negramina essential oil major constituents: ß-myrcene and 2-undocanone, and the transient receptor potential (TRP) channels as potential binding receptors in the aphid and ladybeetles. As the most important results, Negramina essential oil caused mortality in M. persicae aphids with an LC95 = 1.08 mg/cm2, and also significantly repelled the aphids at concentrations as low as 0.14 mg/cm2. Our computational docking analysis reinforced such selectivity actions as the Negramina essential oil major compounds (i.e., ß-myrcene and 2-undocanone) bound to the TRP channels of M. persicae but not to ladybeetle-related TRP channels. Interestingly, the exposure to the Negramina essential oil did not affect the predatory abilities of C. maculata but increased the abilities of E. connexa to prey upon M. persicae. Collectively, our findings provided a physiological basis for the insecticidal and selectivity potential of Negramina essential oil, reinforcing its potential as a tool to be used in integrated pest control programs.

Complete Sequences of Two Plasmids Found in a Brazilian Bacillus thuringiensis Serovar israelensis Strain.

Plasmids play a crucial role in the evolution of bacterial genomes by mediating horizontal gene transfer. In this work, we sequenced two plasmids found in a Brazilian Bacillus thuringiensis serovar israelensis strain which showed 100% nucleotide identities with Bacillus thuringiensis serovar kurstaki plasmids.

Genome Sequence of a New Siphoviridae Phage Found in a Brazilian Bacillus thuringiensis Serovar israelensis Strain.

During the fermentation process, Bacillus thuringiensis (Bt) phages can result in bacterial death and decreased yield. In this work, we describe the genome of a new phage related to the Siphoviridae viral family from a Brazilian strain of Bt which showed high nucleotide sequence identity to the genomes of phages phi4l1 and BtCS33.

Essential oil of Siparuna guianensis as an alternative tool for improved lepidopteran control and resistance management practices.

Although the cultivation of transgenic plants expressing toxins of Bacillus thuringiensis (Bt) represents a successful pest management strategy, the rapid evolution of resistance to Bt plants in several lepidopteran pests has threatened the sustainability of this practice. By exhibiting a favorable safety profile and allowing integration with pest management initiatives, plant essential oils have become relevant pest control alternatives. Here, we assessed the potential of essential oils extracted from a Neotropical plant, Siparuna guianensis Aublet, for improving the control and resistance management of key lepidopteran pests (i.e., Spodoptera frugiperda and Anticarsia gemmatalis). The essential oil exhibited high toxicity against both lepidopteran pest species (including an S. frugiperda strain resistant to Cry1A.105 and Cry2Ab Bt toxins). This high insecticidal activity was associated with necrotic and apoptotic effects revealed by in vitro assays with lepidopteran (but not human) cell lines. Furthermore, deficits in reproduction (e.g., egg-laying deterrence and decreased egg viability), larval development (e.g., feeding inhibition) and locomotion (e.g., individual and grouped larvae walking activities) were recorded for lepidopterans sublethally exposed to the essential oil. Thus, by similarly and efficiently controlling lepidopteran strains susceptible and resistant to Bt toxins, the S. guianensis essential oil represents a promising management tool against key lepidopteran pests.

A recombinant truncated Cry1Ca protein is toxic to lepidopteran insects and forms large cuboidal crystals in insect cells.

A truncated version of the cry1Ca gene from Bacillus thuringiensis was introduced into the genome of Autographa californica multiple nucleopolyhedrovirus (AcMNPV) under the control of two promoters. A recombinant virus (vSyncry1c) was isolated and used to infect insect cells in culture and insect larvae. Structural and ultrastructural analysis of insects infected with vSyncry1C showed the formation of large cuboidal crystals inside the cytoplasm of insect cells in culture and in insect cadavers late in infection. Infected insect cell extracts were analyzed by SDS-PAGE and Western blot and showed the presence of a 65-kDa polypeptide probably corresponding to the protease processed form of the toxin. Bioassays using purified recombinant toxin crystals showed a CL(50) of 19.49 ng/ml for 2(nd) instar A. gemmatalis larvae and 114.1 ng/ml for S. frugiperda.